Stem cankers on sunflower (Helianthus annuus) in Australia reveal a complex of pathogenic Diaporthe (Phomopsis) species.

The identification of Diaporthe (anamorph Phomopsis) species associated with stem canker of sunflower (Helianthus annuus) in Australia was studied using morphology, DNA sequence analysis and pathology. Phylogenetic analysis revealed three clades that did not correspond with known taxa, and these are believed to represent novel species. Diaporthe gulyae sp. nov. is described for isolates that caused a severe stem canker, specifically pale brown to dark brown, irregularly shaped lesions centred at the stem nodes with pith deterioration and mid-stem lodging. This pathogenicity of D. gulyae was confirmed by satisfying Koch's Postulates. These symptoms are almost identical to those of sunflower stem canker caused by D. helianthi that can cause yield reductions of up to 40% in Europe and the USA, although it has not been found in Australia. We show that there has been broad misapplication of the name D. helianthi to many isolates of Diaporthe ( Phomopsis) found causing, or associated with, stem cankers on sunflower. In GenBank, a number of isolates had been identified as D. helianthi, which were accommodated in several clades by molecular phylogenetic analysis. Two less damaging species, D. kochmanii sp. nov. and D. kongii sp. nov., are also described from cankers on sunflower in Australia.

First report of shoot blight of persimmon caused by Diaporthe neotheicola in Australia

Shoot blight symptom was found on persimmon (Diospyros kaki) in southern Western Australia in December 2010. The pathogen was isolated and identified as Diaporthe neotheicola on the basis of morphology, sequence analysis of the internal transcribed spacer (ITS) and the translation elongation factor 1-α (TEF). A pathogenicity test was conducted and Koch's postulates were fulfilled by re-isolation of the fungus from diseased tissues. This is the first report of D. neotheicola causing shoot blight on persimmon in Australia and worldwide. © 2012 Australasian Plant Pathology Society Inc.

Molecular phylogenetic analysis reveals six new species of Diaporthe from Australia

Six new species of Diaporthe, D. beilharziae on Indigofera australis, D. fraxini-angustifoliae on Fraxinus angustifolia subsp. oxycarpa, D. litchicola on Litchi chinensis, D. nothofagi on Nothofagus cunninghamii, D. pascoei on Persea americana and D. salicicola on Salix purpurea from Australia are described and illustrated based on morphological characteristics and molecular analyses. Three of the new species no longer produced sporulating structures in culture and two of these were morphologically described from voucher specimens. Phylogenetic relationships of the new species with other Diaporthe species are revealed by DNA sequence analyses based on the internal transcribed spacer (ITS) region, and partial regions of the β-tubulin (BT) and translation elongation factor 1-alpha (TEF). © 2013 Mushroom Research Foundation.

Green and brown bridges between weeds and crops reveal novel Diaporthe species in Australia

Diaporthe (syn. Phomopsis) species are well-known saprobes, endophytes or pathogens on a range of plants. Several species have wide host ranges and multiple species may sometimes colonise the same host species. This study describes eight novel Diaporthe species isolated from live and/or dead tissue from the broad acre crops lupin, maize, mungbean, soybean and sunflower, and associated weed species in Queensland and New South Wales, as well as the environmental weed bitou bush (Chrysanthemoides monilifera subsp. rotundata) in eastern Australia. The new taxa are differentiated on the basis of morphology and DNA sequence analyses based on the nuclear ribosomal internal transcribed spacer region, and part of the translation elongation factor-1α and ß-tubulin genes. The possible agricultural significance of live weeds and crop residues ('green bridges') as well as dead weeds and crop residues ('brown bridges') in aiding survival of the newly described Diaporthe species is discussed.

Caracterização de Diaporthe citri em diferentes meios de cultura, condições de temperatura e luminosidade

Estudou-se o crescimento micelial de dez isolados de Diaporthe citri, utilizando-se seis meios de cultura (aveia-ágar, maltose-peptona-ágar, batata-dextrose-ágar, folha de laranja-dextrose-ágar, folha de limão-dextrose-ágar, milho-ágar) à temperatura de 22 ± 2 °C e fotoperíodo de 12 h claro/12 h escuro. O cultivo em meio de batata-dextrose-ágar (BDA) foi conduzido em cinco temperaturas diferentes (10, 15, 20, 25 e 30 °C). Três diferentes regimes de luminosidade (12 h claro/12 h escuro, claro contínuo, escuro contínuo) foram utilizados para verificar o crescimento do fungo. Foram observadas variações na produção de picnídios e de massa micelial nos diferentes meios de cultura, temperaturas e regimes de luminosidade testados, sendo que, para a maioria dos isolados, o meio de cultura de aveia-ágar, a faixa de 20 a 25 °C e o regime de claro contínuo induziram maior crescimento micelial. A produção de picnídios foi maior para o regime de luz contínua. O teste de patogenicidade foi feito por inoculação de discos de micélio de 5 mm de diâmetro em ferimentos em ramos e caule de limão 'Feminelo' (Citrus limon) enxertado em citrumelo 'Swingle'(Poncirus trifoliolata x Citrus paradisi) e plantas de limão 'Cravo' (C. limonia) enxertados com laranja 'Valência' (C. sinensis). Após sete dias, houve o aparecimento de exsudação de goma nas plantas inoculadas com os isolados, mas não na testemunha. Todos os isolados mostraram-se patogênicos, sendo os isolados PC2 e PC5, os que causaram comprimento de lesão maior nas plantas.

Caracterização molecular através da técnica fAFLP de isolados de Diaporthe citri

A citricultura é um mercado em expansão, principalmente no Estado de São Paulo, cuja importância na balança comercial já é reconhecida. Como em qualquer espécie cultivada, o crescimento das áreas de cultivo favorecem também o crescimento de problemas fitossanitários. Desta forma, as espécies de citros são afetadas por diversas doenças destacando-se entre elas a melanose, causada por Diaporthe citri (Wolf.), à qual a grande maioria das variedades comerciais são suscetíveis. O conhecimento da diversidade intra-específica é de grande importância, já que esta poderá auxiliar na seleção de variedades com resistência. O objetivo deste trabalho foi avaliar a variabilidade genética em isolados de Diaporthe citri, originários de diferentes locais, variedades e partes da planta, utilizando marcadores moleculares. Marcadores do tipo AFLP (Amplified Fragment Length Polymorphism) foram utilizados para caracterização de dez isolados do patógeno. Os DNAs genômicos extraídos da massa micelial foram utilizados nas reações de amplificação. A técnica fluorescent AFLP permitiu a distinção dos isolados estudados, tendo sido classificados em quatro grupos distintos. Contudo, estes grupos não foram formados em razão da região geográfica, parte da planta ou variedade.

Desenvolvimento de iniciadores para detecção e identificação de Diaporthe phaseolorum var. meridionalis em sementes de soja

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Saprophytic colonization of citrus twigs by Diaporthe citri and factors affecting pycnidial production and conidial survival

Melanose, caused by Diaporthe citri, produces reddish brown lesions on the fruit, leaves, and twigs of citrus trees, and greatly reduces the marketability of fresh fruit. Most of the inoculum is produced in pycnidia on dead twigs in the tree canopy, which exude large numbers of conidia in slimy masses. In this study, detached twigs inoculated with conidia were readily colonized and produced large numbers of pycnidia within 30 to 40 days when they were soaked 3 to 4 h on alternate days. Conidial production was measured by wetting twigs in a rain tower periodically and collecting the conidia in the runoff water. Production began after 80 days and continued for nearly 300 days. In other experiments, production of mature pycnidia on detached twigs was greatest at 94 to 100% relative humidity (RH) and at 28 degrees C. Low RH and temperature, however, favored survival of conidia in exuded masses on twigs. In the field, colonization of detached twigs by D. citri was high in rainy season, moderate in spring and early fall, and minimal in late fall and winter. Twig colonization was positively related to the number of rain days and average temperature, but not to total rainfall. In another experiment, inoculated twigs placed in the tree canopy developed pycnidia and then produced conidial masses for about 200 days. D. citri is a serious pathogen, but a weak parasite, that survives primarily by colonization and reproduction on dead twigs.

Influência de Colletotrichum dematium var. truncata, Phomopsis sojae e Diaporthe phaseolorum f. sp. meridionalis nos testes de vigor de sementes de soja

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Detecção e identificação de Diaporthe phaseolorum var. meridionalis em sementes de soja por PCR

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Fungos endofíticos em Eugenia brasiliensis: prospecção química, biológica, enzimática e avaliação do co-cultivo e epigenética em Xylaria cubensis, Diaporthe sp. e Colletotrichum sp.

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Genetic transformation of Diaporthe phaseolorum, an endophytic fungus found in mangrove forests, mediated by Agrobacterium tumefaciens

We describe the genetic transformation of the mycelial tissue of Diaporthe phaseolorum, an endophytic fungus isolated from the mangrove species Laguncularia racemosa, using Agrobacterium tumefaciens-mediated transformation (ATMT). ATMT uses both the hygromycin B resistant (hph) gene and green fluorescent protein as the selection agents. The T-DNA integration into the fungal genome was assessed by both PCR and Southern blotting. All transformants examined were mitotically stable. An analysis of the T-DNA flanking sequences by thermal asymmetric interlaced PCR (TAIL-PCR) demonstrated that the disrupted genes in the transformants had similarities with conserved domains in proteins involved in antibiotic biosynthesis pathways. A library of 520 transformants was generated, and 31 of these transformants had no antibiotic activity against Staphylococcus aureus, an important human pathogen. The protocol described here, using ATMT in D. phaseolorum, will be useful for the identification and analysis of fungal genes controlling pathogenicity and antibiotic pathways. Moreover, this protocol may be used as a reference for other species in the Diaporthe genus. This is the first report to describe Agrobacterium-mediated transformation of D. phaseolorum as a tool for insertional mutagenesis.

3-Hydroxypropionic Acid as an Antibacterial Agent from Endophytic Fungi Diaporthe phaseolorum

Endophytic fungi are considered a rich source of active compounds resulting from their secondary metabolism. Fungi from marine environment grow in a habitat with unique conditions that can contribute to the activation of metabolic pathways of synthesis of different unknown molecules. The production of these compounds may support the adaptation and survival of the fungi in the marine ecosystem. Mangroves are ecosystems situated between land and sea. They are frequently found in tropical and subtropical areas and enclose approximately 18.1 million hectares of the planet. The great biodiversity found in these ecosystems shows the importance of researching them, including studies regarding new compounds derived from the endophytic fungi that inhabit these ecosystems. 3-hydroxypropionic acid (3-HPA) has been isolated from the mangrove endophytic fungus Diaporthe phaseolorum, which was obtained from branches of Laguncularia racemosa. The structure of this compound was elucidated by spectroscopic methods, mainly 1D and 2D NMR. In bioassays, 3-HPA showed antimicrobial activities against both Staphylococcus aureus and Salmonella typhi. The structure of this antibiotic was modified by the chemical reaction of Fischer-Speier esterification to evaluate the biologic activity of its chemical analog. The esterified product, 3-hydroxypropanoic ethyl ester, did not exhibit antibiotic activity, suggesting that the free carboxylic acid group is important to the pharmacological activity. The antibiotic-producing strain was identified with internal transcribed spacer sequence data. To the best of our knowledge, this is the first report of antibacterial activity by 3-HPA against the growth of medically important pathogens.

Diaporthe aorista; Diaporthaceae

Diaporthe aorista Ellis & Everh.